Effect of human amniotic mesenchymal stem cells transplantation via tail vein on neurological function recovery from ischemic reperfusion injury

doi:10.3969/j.issn.2095-4344.2017.05.009

Abstract

Abstract:

BACKGROUND: Studies have shown that overexpression of myelin-associated glycoprotein (MAG) and oligodendrocyte myelin glycoprotein (OMgp) is the main reason for early neuronal regeneration failure.
OBJECTIVE: To study the effect of human amniotic mesenchymal stem cells (hAMSCs) transplantation on neural functional recovery of rats with ischemia/reperfusion (I/R) injury.
METHODS: Sixty Sprague-Dawley were randomized into sham, I/R, and hAMSCs groups (n=20 per group). Rats in the hAMSCs group were given 1 mL of hAMSCs suspension (1.0×10⁸/L) via the tail vein 24 hours after I/R injury. Rat neurological function recovery was assessed based on behavior changes, as determined by Longa behavioral score, cylinder test, horizontal ladder walking test and limb symmetry test at 1, 3 days, and 1, 2, 3 weeks post transplantation. Cell migration and distribution were observed using immunofluorescence method at 1 day, and 1, 2, 3 weeks post transplantation. MAG and OMgp protein expression was detected by western blot assay at 2 weeks post transplantation. Neuronal apoptosis was detected by TUNEL staining at 3 weeks post transplantation.
RESULTS AND CONCLUSION: In the hAMSCs transplantation group, red marker-positive cells were visible around the injury region at 1 week after transplantation, and over time, these cells were increased in number. Significant improvement in the neurological function of rats were observed in the hAMSCs group as compared with the I/R group at 3 days and 1, 2, 3 weeks after transplantation (P < 0.05), and the expression of MAG and OMgp proteins were also decreased dramatically in the hAMSCs group (P < 0.05). After I/R injury, the number of apoptotic cells was increased, but hAMSCs transplantation reversed this effect. Overall, hAMSCs transplantation can reduce neuronal apoptosis by reducing MAG and OMgp expression levels, and thereby promote neurological functional recovery from I/R injury in rats.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Amnion, Mesenchymal Stem Cell Transplantation, Reperfusion Injury, Tissue Engineering

CLC Number:

R394.2

Zhao Yu. Effect of human amniotic mesenchymal stem cells transplantation via tail vein on neurological function recovery from ischemic reperfusion injury[J]. Chinese Journal of Tissue Engineering Research, 2017, 21(5): 707-712.

Figures/Tables 2

2.1 人羊膜间充质干细胞的形态人羊膜间充质干细胞复苏后重悬于新鲜的细胞培养液中，除少数细胞贴壁死亡外，多数细胞生长良好，培养六七天即可传代，培养14 d可见呈平形排列或漩涡状集落生长，细胞排列紧密(图1A)；第3代细胞生长迅速，纯度高、形态单一(图1B)。实验选取第3代细胞进行移植。 2.2 移植细胞迁移和分布情况免疫荧光检测显示，细胞移植后第1天在损伤区未观察到阳性细胞，移植后1周在损伤区周围可见红色阳性标记细胞，并且随移植时间的延长，移植细胞表达逐渐增多，见图2。 2.3 实验动物数量分析实验共纳入60只SD大鼠，均进入结果分析。 2.4 行为学评分移植后不同时间点进行Longa行为学评分、圆筒实验评分、水平梯行走实验评分、肢体对称性实验评分。结果显示：在移植后24 h各组行为学评分差异无显著性意义；移植后3 d及1，2，3周，细胞移植组较缺血再灌注损伤组神经受损得到明显改善(P < 0.05)；对人羊膜间充质干细胞移植组不同时间点进行比较发现，随着时间的延长，大鼠神经功能亦得到明显改善(P < 0.05)，而缺血再灌注组不同时间点行为学评分比较，差异无显著性意义(P > 0.05)，见表1-4。 2.5 各组大鼠脑组织MAG及OMgp蛋白的表达变化与假手术组比较，缺血再灌注组MAG及OMgp蛋白的表达量均明显升高；与缺血再灌注损伤组比较，人羊膜间充质干细胞移植组MAG及OMgp蛋白的表达量均显著降低，差异有显著性意义(P < 0.05)，见表5和图3。 2.6 TUNEL染色观察神经元凋亡情况与假手术组相比，缺血再灌注损伤组凋亡细胞数增多；人羊膜间充质干细胞移植组凋亡细胞数目较缺血再灌注损伤组下降，见图4。"

References

[1] Jia Q, Liu LP, Wang YJ. Stroke in China. Clin Exp Pharmacol Physiol. 2010;37(2):259-264.

[2] Pandey AK, Bhattacharya P, Shukla SC,et al.Resveratrol inhibits matrix metalloproteinases to attenuate neuronal damage in cerebral ischemia: a molecular docking study exploring possible neuroprotection. Neural Regen Res. 2015; 10(4): 568-575.

[3] Zhao D, Liu J, Wang W, et al. Epidemiological transition of stroke in China: twenty-one-year observational study from the Sino-MONICA-Beijing Project. Stroke. 2008;39(6):1668-1674.

[4] 蒋建平,李静,陈永顺,等.冰七片对大鼠局灶性脑缺血再灌注损伤的保护作用研究[J]. 中外医学研究,2016, 14(1):148-149.

[5] Gao HJ,Liu PF,Li PW,et al.Ligustrazine monomer against cerebral ischemia/reperfusion injury. Neural Regen Res. 2015;10(5): 832-840.

[6] Wu PF, Zhang Z, Wang F, et al. Natural compounds from traditional medicinal herbs in the treatment of cerebral ischemia/reperfusion injury.Acta Pharmacol Sin. 2010;31(12): 1523-1531.

[7] Martino G, Pluchino S, Bonfanti L, et al. Brain regeneration in physiology and pathology: the immune signature driving therapeutic plasticity of neural stem cells. Physiol Rev. 2011; 91(4):1281-1304.

[8] Cafferty WB, Duffy P, Huebner E, et al. MAG and OMgp synergize with Nogo-A to restrict axonal growth and neurological recovery after spinal cord trauma. J Neurosci. 2010;30(20):6825-6837.

[9] Uccelli A, Benvenuto F, Laroni A, et al. Neuroprotective features of mesenchymal stem cells. Best Pract Res Clin Haematol. 2011;24(1):59-64.

[10] Andres RH, Horie N, Slikker W, et al. Human neural stem cells enhance structural plasticity and axonal transport in the ischaemic brain.Brain. 2011;134(Pt 6):1777-1789.

[11] Zhu JM, Zhao YY, Chen SD, et al. Functional recovery after transplantation of neural stem cells modified by brain-derived neurotrophic factor in rats with cerebral ischaemia. J Int Med Res. 2011;39(2):488-498.

[12] Hosseini SM, Farahmandnia M, Razi Z,et al.12 hours after cerebral ischemia is the optimal time for bone marrow mesenchymal stem cell transplantation. Neural Regen Res. 2015;10(6): 904-908.

[13] Ba?enková D, Rosocha J, Tóthová T, et al. Isolation and basic characterization of human term amnion and chorion mesenchymal stromal cells. Cytotherapy. 2011;13(9):1047- 1056.

[14] Lu Y, Hui GZ,Wu ZY,et al.Transformation of human amniotic epithelial cells into neuron-like cells in the microenvironment of traumatic brain injury in vivo and in vitro. Neural Regen Res. 2011; 6(10):744-749.

[15] Díaz-Prado S, Muiños-López E, Hermida-Gómez T, et al. Human amniotic membrane as an alternative source of stem cells for regenerative medicine. Differentiation. 2011;81(3): 162-171.

[16] Huang H, Liu N, Wang JH, et al. The effects of adipose-derived stem cells transplantation on the expression of TGF-β1 in rat brain after cerebral ischemia. Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2011;27(8):872-875.

[17] Sapin V, Souteyrand G, Bonnin N, et al. Therapeutic use of amniotic membranes and their derivate cells. Gynecol Obstet Fertil. 2011;39(6):388-390.

[18] Sun F, Xie L, Mao X, et al. Effect of a contralateral lesion on neurological recovery from stroke in rats. Restor Neurol Neurosci. 2012;30(6):491-495.

[19] Pignataro G, Meller R, Inoue K, et al. In vivo and in vitro characterization of a novel neuroprotective strategy for stroke: ischemic postconditioning. J Cereb Blood Flow Metab. 2008; 28(2):232-241.

[20] Zhang Q, Yuan W, Wang G, et al. The protective effects of a phosphodiesterase 5 inhibitor, sildenafil, on postresuscitation cardiac dysfunction of cardiac arrest: metabolic evidence from microdialysis. Crit Care. 2014;18(6):641.

[21] 王灿,王琳琳,苗明三.脑脉舒康胶囊对沙土鼠脑缺血再灌注损伤的影响[J].中华中医药杂志,2014,29(7):2352-2355.

[22] Miao MS,Guo L,Li RQ,et al.Radix Ilicis Pubescentis total flavonoids combined with mobilization of bone marrow stem cells to protect against cerebral ischemia/reperfusion injury. Neural Regen Res. 2016;11(2): 278-284.

[23] 刘灵芝.巴戟天醇提物对离体大鼠缺血再灌注损伤心肌能量代谢的影响[J].中国医药导报,2012,9(28):10-11。

[24] 张向前,王雪侠.巴戟天醇提物对大鼠肾缺血再灌注损伤的保护作用[J].中国医药导报,2013,10(5):22-24.

[25] 赵刚,王伟民,杨帅,等.白黎芦醇对大鼠脑缺血再灌注损伤的保护作用[J].中国老年学杂志,2014,27(21):6149-6150.

[26] 肖爱娇,陈日新,康明非,等.热敏灸对脑缺血再灌注损伤大鼠SOD、MDA的影响[J].天津医药,2014,27(1):51-53.

[27] Peng B, Guo QL, He ZJ, et al. Remote ischemic postconditioning protects the brain from global cerebral ischemia/reperfusion injury by up-regulating endothelial nitric oxide synthase through the PI3K/Akt pathway. Brain Res. 2012;1445:92-102.

[28] Wang WB,Yang LF,He QS,et al.Mechanisms of electroacupuncture effects on acute cerebral ischemia/ reperfusion injury: possible association with upregulation of transforming growth factor beta 1. Neural Regen Res. 2016; 11(7): 1099-1101.

[29] Xie D, Zhang EL, Li JM, et al. Design, synthesis and anti-platelet aggregation activities of ligustrazine- tetrahydroisoquinoline derivatives. Yao Xue Xue Bao. 2015; 50(3):326-331.

[30] Hu J, Lang Y, Cao Y, et al. The Neuroprotective Effect of Tetramethylpyrazine Against Contusive Spinal Cord Injury by Activating PGC-1α in Rats. Neurochem Res. 2015;40(7): 1393-1401.

[31] Li N, Deng XG, Zhang SH, et al. Effects of different concentrations of tetramethylpyrazine, an active constituent of Chinese herb, on human corneal epithelial cell damaged by hydrogen peroxide. Int J Ophthalmol. 2014;7(6):947-951.

[32] 马淑媛,高芳.川穹嗪注射液治疗急性脑梗死临床疗效观察[J].牡丹江医学院学报,2008, 29(6): 34-35.

[33] 余奇劲,杨洁,陈娟. 缺血前和缺血后联合应用丙泊酚对兔脊髓缺血再灌注损伤的保护作用[J].中华临床医师杂志,2011, 5(20): 5955-5959.

[34] Moon JH, Lee JR, Jee BC, et al. Successful vitrification of human amnion-derived mesenchymal stem cells. Hum Reprod. 2008;23(8):1760-1770.

[35] Lei L,Zhou RX. Migration and differentiation of bone marrow-derived multipotent adult progenitor cells through tail vein injection in a rat model of cerebral ischemia. Neural Regen Res. 2009; 4(2):118-122.

[36] Pan FH, Ding XS, Ding HX, et al. Stem cell transplantation for treatment of cerebral ischemia in rats: effects of human umbilical cord blood stem cells and human neural stem cells. Neural Regen Res. 2010; 5(7):485-490.

[37] 郝永岗,张正春,薛群,等.骨髓间充质干细胞移植治疗脑出血的实验研究[J].中国急救医学,2009,29(7):615-618.